PROJECT SUMMARY (ABSTRACT): Background: TET1, the founding member of the TET methylcytosine dioxygenase family, was first identified as a fusion partner of the MLL gene in acute myeloid leukemia (AML). In contrast to the previous thought that all three TET genes (TET1/2/3) may function as tumor suppressor genes in cancers, our recent work showed that TET1 is overexpressed in human MLL-rearranged AML and also AML subtypes carrying t(8;21), FLT3-ITD and/or NPM1 mutations, and plays a critical oncogenic role in the development of such AMLs. However, the role of TET1 in the pathogenesis of other types of hematopoietic malignancies, such as myelodysplastic syndromes (MDS), remains unclear. Very recently, in analysis of a genome-wide gene expression profiling dataset of a large cohort of human primary MDS patients, we found that TET1 is also aberrantly overexpressed in all the major subtypes of MDS analyzed. We showed that knockdown of TET1 expression substantially inhibited the growth/proliferation and colony-forming ability of human MDS cells, and the opposite is true when TET1 is forced expressed. In addition, we have recently developed effective small- molecule compounds (i.e., NSC-370284 and its more effective structural analog UC-514321) that selectively target STAT/TET1 signaling and exhibit potent therapeutic efficiency in treating AML. Remarkably, we showed that UC-514321 also exhibits potent anti-MDS effects in vitro and in vivo. Objective/Hypothesis: TET1 plays an essential role in MDS pathogenesis through regulating expression of a set of essential target genes and that targeting TET1 signaling is a promising therapeutic strategy to treat MDS. Specific Aims: (1) To determine the pathological function of TET1 in the development and maintenance of MDS; (2) To assess the therapeutic potential of targeting TET1 signaling to treat human MDS; and (3) To identify critical target genes of TET1 in MDS. Study Design: 1) We will conduct both gain- and loss-of-function studies of TET1/Tet1 in a series of MDS animal models to define the role of TET1/Tet1 in both development and maintenance of MDS (Aim 1). 2) We will use in vivo patient-derived xeno-transplantation (PDX) MDS models to determine whether targeting TET1 signaling by small-molecule FTO inhibitors (e.g., UC-514321) alone and especially together with other therapeutic agents (e.g., decitabine; DAC) is an effective therapeutic strategy to treat human MDS. The therapeutic effects will also be assessed in various murine MDS models in vivo (Aim 2). 3) We will perform genome-wide ChIP-seq, 5hmC-seq, and RNA-seq to identify critical direct targets of TET1/Tet1, followed by the validation and functional studies of a group of selected TET1/Tet1 targets in vitro and in vivo, to elucidate the underlying molecular mechanism(s) (Aim 3).